title: Comparison of fluorescence reagents for simultaneous determination of hydroxylated phenylalanine and nitrated tyrosine by high-performance liquid chromatography with fluorescence detection.
abstract: Reactive oxygen species (ROS) and reactive nitrogen species (RNS) are well-known and important contributors to oxidative and nitrosative stress in several diseases. Hydroxylated phenylalanine and nitrated tyrosine products appear to be particularly susceptible targets of oxidative and nitrosative stress. We compared fluorescence reagents for their potential use in the analysis of hydroxylated phenylalanine and nitrated tyrosine products with a high-sensitivity and high-specificity HPLC-UV-FL technique. The analytes were extracted from serum via solid-phase extraction on Waters Oasis MCX cartridges. Chromatographic separation was achieved on an ODS column (Capcell Pak MG II; 150 × 2.0 mm) using a gradient mobile phase consisting of 20 mm sodium phosphate buffer (adjusted to pH 3.0) and acetonitrile. The method quantification limit for 4-nitrophenylalanine, m-tyrosine, and 3-nitrotyrosine was 0.1 μm. The relative standard deviation of the precision and accuracy was acceptable at the spiked concentration of 0.1 μm for 4-nitrophenylalanine, m-tyrosine and 3-nitrotyrosine. The method could be used for the in vitro analysis of serum samples.
authors.affiliation: Department of Analytical Chemistry, Faculty of Pharmaceutical Sciences, Hoshi University, Tokyo, Japan.
authors.forename: Yusuke
authors.initials: Y
authors.lastname: Iwasaki
authors.forename: Keisuke
authors.initials: K
authors.lastname: Mochizuki
authors.forename: Yuki
authors.initials: Y
authors.lastname: Nakano
authors.forename: Natsumi
authors.initials: N
authors.lastname: Maruya
authors.forename: Masato
authors.initials: M
authors.lastname: Goto
authors.forename: Yosuke
authors.initials: Y
authors.lastname: Maruyama
authors.forename: Rie
authors.initials: R
authors.lastname: Ito
authors.forename: Koichi
authors.initials: K
authors.lastname: Saito
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Nakazawa
mesh_terms: D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D005456:Fluorescent Dyes / Q000032:analysis* / Q000737:chemistry; D006801:Humans; D006861:Hydrogen Peroxide; D006863:Hydrogen-Ion Concentration; D057230:Limit of Detection; D030421:Peroxynitrous Acid; D010649:Phenylalanine / Q000031:analogs & derivatives* / Q000032:analysis; D015203:Reproducibility of Results; D052616:Solid Phase Extraction; D013050:Spectrometry, Fluorescence / Q000379:methods* / Q000592:standards; D014443:Tyrosine / Q000031:analogs & derivatives* / Q000032:analysis
chemical_list: D005456:Fluorescent Dyes; D030421:Peroxynitrous Acid; C040239:4-nitrophenylalanine; C002744:3-nitrotyrosine; D014443:Tyrosine; D010649:Phenylalanine; D006861:Hydrogen Peroxide; C015566:3-tyrosine